Phenotypic diversity and metabolic specialization of renal endothelial cells

Abstract

Complex multicellular life in mammals relies on functional cooperation of different organs for the survival of the whole organism. The kidneys play a critical part in this process through the maintenance of fluid volume and composition homeostasis, which enables other organs to fulfil their tasks. The renal endothelium exhibits phenotypic and molecular traits that distinguish it from endothelia of other organs. Moreover, the adult kidney vasculature comprises diverse populations of mostly quiescent, but not metabolically inactive, endothelial cells (ECs) that reside within the kidney glomeruli, cortex and medulla. Each of these populations supports specific functions, for example, in the filtration of blood plasma, the reabsorption and secretion of water and solutes, and the concentration of urine. Transcriptional profiling of these diverse EC populations suggests they have adapted to local microenvironmental conditions (hypoxia, shear stress, hyperosmolarity), enabling them to support kidney functions. Exposure of ECs to microenvironment-derived angiogenic factors affects their metabolism, and sustains kidney development and homeostasis, whereas EC-derived angiocrine factors preserve distinct microenvironment niches. In the context of kidney disease, renal ECs show alteration in their metabolism and phenotype in response to pathological changes in the local microenvironment, further promoting kidney dysfunction. Understanding the diversity and specialization of kidney ECs could provide new avenues for the treatment of kidney diseases and kidney regeneration.

Fig. 1. Anatomy and heterogeneity of the kidney vasculature.

a | The human kidney contains a highly specialized vasculature that is closely linked to the kidney epithelial system. The blood vascular system delivers oxygen and nutrients to parenchymal tissues, and facilitates waste removal, immune surveillance and immune cell trafficking, coagulation, and the production of angiocrine signals for tissue maintenance and regeneration, whereas the lymphatic vascular system drains the extravasated interstitial fluid from permeable blood capillaries back to the veins, and facilitates immune cell trafficking and lipid transport. b | Endothelial cells (ECs) are characterized by their tissue-specific transcriptional heterogeneity. The plot shows a representative dimensionality reduction of the transcriptome profiles of ECs isolated from kidney and four other hypothetical tissues or organs. c | EC heterogeneity within the kidney is dictated by a combination of genetic factors and exposure to different microenvironments, which ultimately contribute to kidney function.

Fig. 2. Phenotypic and molecular heterogeneity of the glomerular endothelium.

a | The glomerular vasculature comprises the glomerular capillaries connected to the afferent and efferent arterioles. The arterioles are in partial contact with the juxtaglomerular apparatus (JGA), which comprises the macula densa of the distal convoluted tubule, granular renin-producing cells and extraglomerular mesangial cells. b | The glomerular filtration barrier is composed of glomerular capillary endothelial cells (ECs) and podocytes, separated by a glomerular basement membrane (GBM). The glomerular capillary endothelium encompasses non-diaphragmed fenestrations and a thick glycocalyx, which sustain glomerular filtration and permselectivity. c | The heterogeneity of glomerular renal ECs (gRECs) is demonstrated by the differential expression of genes by different gREC types. Pink and purple reflect arteriole and capillary EC phenotypes, respectively. Since REC subpopulations express a combination of several markers, these are indicated following a hierarchical system. More detailed information can be found in Supplementary Table 1. d | The layer of vascular smooth muscle cells from the afferent arteriole is progressively replaced by renin-expressing granular cells within the JGA at the entrance of the glomerular tuft. A switch from a non-fenestrated and Cxcr4-negative endothelium to a fenestrated and Cxcr4-positive endothelium within the JGA is also observed.

Fig. 3. Phenotypic and molecular heterogeneity of the cortical and medullary renal endothelium.

a | Phenotypically distinct renal endothelial cell (REC) phenotypes coexist within the two main anatomical compartments of the kidney, the cortex and medulla. b | Markers of different cortical REC (cREC) phenotypes. Since REC subpopulations express a combination of several markers, these are indicated following a hierarchical system. c | Markers of different medullary REC (mREC) populations. Since REC subpopulations express a combination of several markers, these are indicated following a hierarchical system. More detailed information regarding the expression and function of genes expressed in cortical and medullary RECs can be found in Supplementary Table 1. d | Phenotypic differences exist between the descending vasa recta (DVR) and the ascending vasa recta (AVR). The arterial-like ECs of the DVR are non-fenestrated and covered by a pericyte layer that regulates the medullary blood flow. By contrast, the venous-like ECs of the AVR are highly fenestrated and lack pericyte coverage, which facilitates water reuptake.

Fig. 4. Exposure of the renal endothelium to changes in oxygen tension.

a | The kidney is exposed to hypoxia under both physiological and pathological circumstances, such as in acute kidney injury (AKI) and chronic kidney disease (CKD). b | Endothelial cells (ECs) adapt to hypoxia by stabilizing and activating the transcription factor hypoxia inducible factor (HIF) through the direct effects of low O₂ levels, the release of ATP and downstream activation of the adenosine A2a receptors, the production of NADPH-derived and oxidative phosphorylation (OXPHOS)-derived reactive oxygen species (ROS), and as a consequence of sirtuin 3 (SIRT3) activation and the upregulation or activation of fatty acid synthase (FAS) (left panel). Exposure to hypoxia affects the metabolism of ECs by enhancing anaerobic metabolism and fatty acid biosynthesis while repressing aerobic metabolism (OXPHOS). Levels of the metalloenzyme arginase II are increased whereas arginine uptake is repressed, leading to low intracellular arginine availability, which results in endothelial nitric oxide synthase (eNOS) uncoupling and the production of ROS (central panel). ECs counteract ROS produced as a result of the exposure to hypoxia by activating a SIRT3–forkhead box 3 (FOXO3)-dependent antioxidant gene expression pathway (right panel). CAT, cationic amino acid transporter; endoMT, endothelial-to-mesenchymal transition; GLUT, glucose transporter; LAT, large neutral amino acid transporter.

Fig. 5. Response of the renal endothelium to changes in blood flow and shear stress.

a | Arteries, arterioles, capillaries, venules and veins are exposed to different types of blood flow as defined by its pulsatility and the level of shear stress. b | Glomerular capillaries are exposed to high shear stress as a result of high blood flow and pressure together with increasing blood viscosity due to the filtration process. c | Some regions of renal arteries and arterioles — which control kidney blood perfusion and glomerular filtration rate — are exposed to laminar blood flow, whereas other regions — particularly regions of vessel bifurcation — are exposed to disturbed blood flow. Regions of disturbed blood flow are more likely to develop atherosclerotic lesions. d | Endothelial cells (ECs) exposed to laminar blood flow inhibit glycolytic metabolism in a KLF2-dependent manner and divert glycolytic intermediates to pathways involved in glycocalyx biosynthesis (left panel). Laminar flow also induces SIRT1–PGC1α–TFAM-mediated mitochondrial biogenesis, and stimulates the production of oxidative phosphorylation (OXPHOS)-derived ATP, which activates purinergic P2X4 receptors and further induces the activation of KLF2 and KLF4 transcription factors. Moreover, Krüppel-like factor 2 (KLF2) and KLF4 are upregulated in response to MEK5–ERK5 pathway activation (right panel). Laminar blood flow promotes EC quiescence, the production of antioxidants, the formation of a tight endothelial barrier and the acquisition of an anticoagulant and anti-inflammatory phenotype, and protects against proteinuria while maintaining glomerular filtration. e | Disturbed blood flow represses the protective pathways induced by laminar blood flow. Induction of microRNA (miR)-92a by disturbed flow represses the endothelial expression of KLF2, KLF4 and SIRT1 and PPAP2B, releasing the inhibitory effect of PPAP2B on circulating lysophosphatidic acid (LPA). Binding of LPA to its receptor LPAR1 induces pro-inflammatory signalling. The induction of SREBP2 upregulates HMG-CoA reductase, which increases intracellular cholesterol levels, and further upregulates miR-92a. Hypermethylation of the KLF4 promoter prevents MEF2 binding and subsequent KLF4 transcription (left panel). ECs exposed to disturbed blood flow also demonstrate an uncoupling of glycolysis from mitochondrial metabolism, with an upregulation of glycolysis. Disturbed flow also induces the production of NOX4-derived reactive oxygen species (ROS) and activates nuclear factor κB (NF-κB), leading to the upregulation and stabilization of hypoxia inducible factor 1α (HIF1α). Activation of the NF-κB pathway has been linked to heparanase activity and consequent glycocalyx degradation (right panel). Disturbed blood flow ultimately triggers EC activation and oxidative stress, impairs the endothelial barrier, induces a procoagulant and pro-inflammatory EC phenotype, and favours the development of atherosclerotic renal artery stenosis. ERK, extracellular signal regulated kinase; G6P, glucose-6-phosphate; GLUT, glucose transporter; HA, hyaluronan; HAS2, hyaluronan synthase 2; HMGCoAR, hydroxymethylglutaryl CoA reductase; MEF2, myocyte enhancer factor-2; MEK, mitogen-activated protein kinase kinase; NOX4, NADPH oxidase 4; PDK1, pyruvate dehydrogenase kinase 1; PGC1α, peroxisome proliferator-activated receptor gamma coactivator-1α; PPAP2B, phosphatidic acid phosphatase type 2B; SIRT1, sirtuin 1; SREBP2, sterol regulatory element binding protein 2; TFAM, mitochondrial transcription factor A; UDP-GlcNAc, uridine diphosphate-N-acetylglucosamine; UDP-GlcUA, uridine diphosphate–glucuronic acid.

Fig. 6. Response of the renal endothelium to changes in osmolarity.

a | The renal medullary and papillary regions of the kidney are exposed to hyperosmolarity as a consequence of the countercurrent multiplier and exchange mechanisms, which generates and maintains the medullary hyperosmolarity gradient (ranging from 300 mOsm/kg H₂O at the corticomedullary junction to up to 1,400 mOsm in the papilla) that drives the process of urine concentration. b | In response to a rapid increase in osmolarity (for example, following a switch from diuresis to anti-diuresis) endothelial cells (ECs) tend to shrink as a consequence of water loss. This response results in cytoskeletal rearrangements and activation of a regulatory volume increase (RVI) compensatory mechanism, characterized by an accumulation of intracellular Na⁺ and urea followed by osmotic water reabsorption. Moreover, the expression of heat shock proteins is induced to preserve the correct folding of proteins from high levels of denaturing urea (left panel). Prolonged exposure to hyperosmolarity, such as occurs in the papilla or during situations of prolonged dehydration, induces ECs to promote the production of ATP from oxidative phosphorylation (OXPHOS), and stimulate active Na⁺ export through the Na⁺/K⁺ ATPase as well as the import and synthesis of inert organic osmolytes (such as glucose-derived polyols) to protect the cell from hyperosmolarity-induced cell damage (right panel). AVR, ascending vasa recta; DVR, descending vasa recta; GLUT, glucose transporter; ROS, reactive oxygen species.

Fig. 7. Sprouting and intussusceptive angiogenesis.

a | Sprouting angiogenesis occurs after a stimulus with an angiogenic growth factor such as VEGF, which activates endothelial cells on pre-existing blood vessels. The activated endothelial cells (ECs), called tip cells, release enzymes that degrade the basement membrane to allow the ECs to migrate from the pre-existing blood vessel, initiating the sprout. The endothelial cells that follow the tip cells (called stalk cells) proliferate to enable extension of the sprout towards the angiogenic stimulus. When two tip cells meet they fuse to form a new capillary lumen that undergoes further vessel maturation and stabilization. b | Intussusceptive angiogenesis, also called splitting angiogenesis, occurs by splitting a pre-existing blood vessel into two. This process begins with the formation of a pillar extension that protrudes towards the vessel lumen, and forms a transcapillary pillar that splits the vessel into two. Concurrently, myofibroblasts migrate towards the new pillar to help stabilize the newly formed vessels through the deposition of collagen fibres.